Toxoplasmic encephalitis is a major opportunistic infection in AIDS patients. Many of these patients cannot tolerate prolonged administration of pyrimethamine-sulfadiazine. Thus new drugs are needed to treat to prevent this disease. The rational search for improved drug would profit from knowledge as to the mechanism of action of current and future candidate drugs. A useful clue as to mechanism of action can be obtained by studying drug-resistant mutants of Toxoplasma gondii. One way to take advantage of these mutants is to use genetic analysis to identify the gene that confers resistance. Now that is a formal genetic map of T. gondii based on RFLP markers has been established, it is possible to localize drug-resistance genes. This Core Facility will isolate drug-resistant mutants of the PLK strain of T. gondii with the aid of chemical mutagenesis. These mutants will be crossed with a CEP strains of T. gondii that has three unlinked drug-resistance markets. These crosses will be carried out in the definitive host, the cat. Oocysts from the cat feces will be purified and allowed to sporulate. The bradyzoites will be released from sporulated oocysts and grown as tachyzoites in cultured human fibroblasts. Recombinant parasites that inherited drug resistance markers from both parents will be cloned and characterized. The recombinant clones will be sent to Dr. John Boothroyd's laboratory or RFLP analysis.